Abstract
BackgroundSerum-free xenogen-free defined media and continuous controlled physiological cell culture conditions have been developed for stem cell therapeutics, but the effect of these conditions on the relative potency of the cell product is unknown. As such, we conducted a head-to-head comparison of cell culture conditions on human heart explant-derived cells using established in vitro measures of cell potency and in vivo functional repair.MethodsHeart explant-derived cells cultured from human atrial or ventricular biopsies within a serum-free xenogen-free media and a continuous physiological culture environment were compared to cells cultured under traditional (high serum) cell culture conditions in a standard clean room facility.ResultsTransitioning from traditional high serum cell culture conditions to serum-free xenogen-free conditions had no effect on cell culture yields but provided a smaller, more homogenous, cell product with only minor antigenic changes. Culture within continuous physiologic conditions markedly boosted cell proliferation while increasing the expression of stem cell-related antigens and ability of cells to stimulate angiogenesis. Intramyocardial injection of physiologic cultured cells into immunodeficient mice 1 week after coronary ligation translated into improved cardiac function and reduced scar burden which was attributable to increased production of pro-healing cytokines, extracellular vesicles, and microRNAs.ConclusionsContinuous physiological cell culture increased cell growth, paracrine output, and treatment outcomes to provide the greatest functional benefit after experimental myocardial infarction.
Highlights
Serum-free xenogen-free defined media and continuous controlled physiological cell culture conditions have been developed for stem cell therapeutics, but the effect of these conditions on the relative potency of the cell product is unknown
Primary Explant-derived cell (EDC) cultures were established by plating half of each atrial appendage specimen in standard serum-supplemented media and half in serum-free xeno-free medium (SF) medium (50:50 split by mass, Fig. 1a) within standalone oxygen-controlled incubators and level 2 biosafety cabinets in a standard clean room facility (STD env, Fig. 1b)
In contrast to previous work demonstrating the effect of divergent culture practices on EDC biology [12,13,14], Good manufacturing practices (GMP) grade collagenase did not influence either overall cell culture yields (20 ± 5 versus 21 ± 6 × 106 cells per mg tissue plated, respectively; p = 0.92) or the major sub-population content at each serial harvest from the plated tissue (Additional file 1: Figure S1)
Summary
Serum-free xenogen-free defined media and continuous controlled physiological cell culture conditions have been developed for stem cell therapeutics, but the effect of these conditions on the relative potency of the cell product is unknown. We conducted a head-to-head comparison of cell culture conditions on human heart explant-derived cells using established in vitro measures of cell potency and in vivo functional repair. Health care systems are faced with a growing number of patients living with chronic heart failure—a diagnosis that still carries a 5-year mortality approaching 50% [1, 2]. EDCs are considerably more apt to grow into working heart tissue than cardiospherederived cells, clinical delivery has not been possible as the human cell “dose,” estimated to be 25–75 M based on cardiosphere-derived cell trials, greatly exceeds production capacity using traditional methods [9]. We focused on cell production, an important and often understudied area of research, to identify novel means of enhancing EDC cell culture outcomes
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